Technical Field
The present disclosure relates to solid phase extraction techniques of molecules of interest, and apparatus for achieving these techniques. The techniques of the present disclosure eliminate the need for centrifugation of samples standard in conventional solid phase extraction. The techniques of the present disclosure also eliminate the need for specialized laboratory skills and equipment (e.g. micropipettes).
More specifically, the present disclosure relates to apparatus for passing a volume of sample using a syringe device through sorbent to bind a substance of interest and subsequently eluting the substance of interest from the sorbent using the syringe device without the need for centrifugation. In illustrative embodiments, examples of sorbents for isolating nucleic acids as the substance of interest include, but are not limited to: silica, acid washed silica, glass beads, acid washed glass beads, zeolite, silica gel, filters embedded with silica particles, or mixtures of the above.
Description of Related Art
Solid phase extraction (SPE) is a useful sample preparation technique for isolating or purifying compounds of interest from a liquid sample matrix. SPE relies on physical properties of compounds of interest to capture them from the sample matrix. For example, SPE is used to isolate DNA from cells lysate, whole blood, serum, plasma, sputum, urine, fecal, semen, cerebro-spinal fluid or oral fluids. SPE is an efficient process, and advantageously avoids problems common in other liquid/liquid extraction techniques such as incomplete phase separations, specialty glassware and the need to use and dispose large quantities of organic solvents.
SPE samples are frequently prepared for downstream use in polymerase chain reaction (PCR), where contaminants or other molecules not of interest to the analysis could inhibit the PCR. Other molecular diagnostic testing modalities exist, such as NASBA, RPA, HDA, LAMP, RCA, ICAN, SMART, SDA, microarray, and LDR. All molecular diagnostic testing modalities benefit from a stronger signal through sample isolate purity. The purities achievable using SPE render it an attractive tool for molecular diagnostics for public health laboratories, biotechnology companies, government agencies, law enforcement agencies, and research institutes.
There are numerous technique variations for using SPE, which vary by how compounds are retained by a substance known as the sorbent. Variants include reversed-phase, normal-phase, ion-exchange and adsorption. A common approach that uses a silica sorbent membrane is the spin column procedure known as QIAamp available from Qiagen N.V., Venlo, The Netherlands, used for instance for nucleic acid purification from tissues, swabs or body fluids (e.g., blood, sweat, washed urine cells, semen, cerebro-spinal fluid, etc). In this procedure, depicted in FIG. 1 and discussed in greater detail below, samples are lysed, binded to the silica membrane and washed. The purified nucleic acid is then eluted from the silica membrane.
Unfortunately, the prior art approach for sample extraction involves either the use of micropipettes and centrifuges to process the sample at each stage. Further, the volume of the sample in traditional extraction is typically limited, for example to 140 μl, and greater volumes require additional centrifugations. Traditional silica column based extractions (such as Qiagen, QIAamp Viral RNA, catalog 52904), require that the sample with lysis buffer and alcohol is loaded into the spin column. Based on the size of the spin column, only 630 μL can typically be loaded at one time A 140 μL sample requires 560 μL of lysis buffer and 560 μL of ethanol, which requires two centrifugation spins to load all of the sample/buffer through the silica.